Neonatal calf diarrhea remains one of the most important problems faced by livestock, causing great economic losses. Fecal samples were collected from 100 diarrheic calves in Al-Fayoum governorate, Egypt during 2021, and 2022, to investigate the prevalence of Enterotoxigenic Escherichia coli (ETEC), Salmonella typhimurium, Klebsiella pneumonia, Clostridium perfringens, bovine rotavirus, bovine coronavirus, and Cryptosporidium parvum which are the major enteropathogens associated with neonatal calf diarrhea, the prevalence of enteropathogens were 58%, 29%, 34%, 14%, 35%, 8%, and 65% respectively. Molecular characterization was performed to confirm the E. coli, Klebsiella pneumonia, Salmonella typhimurium, Clostridium perfringens, and Cryptosporidium isolates and to detect some virulent genes associated with their pathogenicity. All the bacterial isolates gave a clear band with 16S rRNA. In E. coli, virulent genes (K99, F41, phoA) were detected, also; Salmonella strains were found positive for the invA and sopB gene, while all Clostridium perfringens strains were tested positive for Alpha and Beta toxin but negative for Epsilon toxin. On the other hand, all Klebsiella pneumoniae isolates were tested positive with iutA and fimH genes. Also, the in-vitro antibiotic sensitivity testing of bacterial isolates was applied. Statistical analysis was carried out to determine the potential influence of age factor on the reported prevalences of concurrent infections, which revealed that the animals age significantly affected the infection prevalence in all pathogens inversely excepts those infected by Klebsiella was affected by age directly, and those infected by E. Coli, or Cryptosporidium, were not affected by age at all. Good hygienic management and good vaccination program are very important to overcome acute diarrhea in neonate calves and the misuse of antibiotic revealed the presence of multidrug resistance isolates of some enteropathogenic bacteria.

Rotavirus ribonucleic acid was extracted from 16 fecal samples of the serologically positive diarrheic calves using Latex agglutination test (LAT) and Immunochrmatographic assay (ICA). The extracted RNA was submitted to Reverse transcriptase polymerase chain reaction (RT-PCR) to detect VP7 and VP4 genes and the positive samples were 100% (16/16) and 81.25% (13/16), respectively. The amplified products were subjected to G and P-genotyping by semi-nested multiplex PCR using of G6, G8 and G10 genotyping and P1, P5 and P11 genotyping primers, respectively. G6 was detected in 10 (62.50%) of 16 samples and G10 was diagnosed in 5 (31.25%) of 16 samples and one (6.25%) sample did not react with any G primer used. P5 was detected in 9 (56.25%) of 16 samples, P11 was diagnosed in 3 (18.75%) of 16 samples, mixed infection with P5+P11 was observed in 1 (6.25%) of 16 samples and 3 (18.75%) samples did not react with any P primer used. G and P genotypes combination revealed that G6P5 was in 50% (8/16), G10P11 in 12.50% (2/16), G10P5 in 6.25% (1/16), G6P11 in 6.25% (1/16), G10 (P5+P11) in 6.25% (1/16), G6P? in 6.25% (1/16), G10P? in 6.25% (1/16), and G?P? in 6.25% (1/16). These results suggest that the detected genotypes can used as dominant strains for the formulation of an appropriate vaccine against BRV in Assiut Governorate. In conclusion, RT-PCR and Semi-nested multiplex PCR can used as rapid and confirmatory test for detection of nucleic acid and genotypes of Rotavirus, G and P genotypes combination in the present study revealed that G6P5, G6P11, G10P5 and G10P11 were circulating genotypes in bovine population in Assiut governorate. G6P5 strain was the most common of all strain diagnosed in other fecal samples. The presence of various combinations of G and P genotypes among field isolates of BRV suggests that genetic reassortment frequently occurred between viral strains with genes encoding different G and P genotypes. Finally, presence of different genotypes of Rotaviruses emphasizes their simultaneous monitoring in animals for the development and optimization of Rotavirus vaccines.

Throughout the world, rotavirus infections cause extensive morbidity in human infants and diarrhea in animals such as white scour caused by bovine rotavirus in calves. We isolated three rotavirus strains designated KV0407, KV0418, and KV0426 from 103 fecal samples of diarrheic calves. The genes coding for proteins VP4, VP6, VP7, and NSP4 from strain KV0407 were sequenced and compared with the nucleotide sequences of other known strains of rotavirus. The KV0407 VP4 gene was highly homologous to the OSU (99.4%) and JL94 (99.4%), but not the B223 (62.4%) and K33 (62.4%) VP4 genes. The KV0407 and KV0418 VP7 genes were most similar to the OSU and super-short type VMRI VP7 genes. Based on nucleotide sequence analysis, the KV0407 strain was tentatively assigned to A serogroup (SG I), G5P[7], NSP4 genotype B and the KV0418 and KV0426 strains were assigned to A serogroup (SG I), G6P[5], NSP4 genotype A. The genetic characterization of these bovine rotavirus isolates could be useful for the diagnosis and prevention of diarrhea in calves.